About 1-3% of all women undergo early menopause, either never going through menarche or stopping menstruation by the mid-30's, rather than reaching the standard reproductive lifespan of about 50. A fraction of such instances of early-onset "premature ovarian failure (POF)" is genetic. A single locus on chromosome 3 is implicated in several families. In collaboration with the group of Dr. G. Pilia, we isolated the gene, FOXL2, in which mutations cause POF and an eyelid malformation (constituting the syndrome Blepharophimosis/Ptosis/Epicanthus Inversus, or BPES). We are now analyzing the function of FOXL2, which appears to determine the level of ovarian follicles. We recovered the mouse orthologue, Foxl2, and generated a knockout mouse model. The resulting phenotype mimics eyelid and ovarian features seen in BPES. In the ovaries of the knockout mice, pervasive failure of follicle formation occurs. Complete individual follicles never form from primitive oocyte nests; development of all somatic lineages in the ovary is blocked; and all oocytes can subsequently be derepressed for growth and undergo apoptosis. The results point toward a comparable overall mechanism for POF in women deficient in FOXL2. In extended studies, we found that the arrest of ovary development is succeeded by a phase of partial sex reversal. To help understand the failure of follicle formation and maintenance in POF at a molecular level, we have analyzed normal ovarian development in mice using the NIA microarrays [developed by the group of Dr. M. Ko (LG)], first with 15K rrays and now with 44K arrays. We have thus far compared the profile of genes expressed in ovaries of newborn mice, in which ovarian follicles are just starting to form, to the cohort of genes expressed in mature ovary, in which follicles are fully formed. Marker genes expressed in follicle cells only at birth, or in oocytes of nascent or in mature mice, have been recovered for further functional studies. These are analyzed in comparison to the subset of genes whose function is sharply affected in hereditary POF, starting with expression profiling of developing ovaries from Foxl2 knockout mice. In that model, the interruption of programmed and coordinated gene function is characterized by determining the target genes regulated by Foxl2. To further the analysis of a possible second role of Foxl2 as a "repressor of maleness", additional mouse models are being constructed that would force expression of Foxl2 at times and in cells that would otherwise become testis- rather than ovary-specific; those models will be studied by careful phenotyping and repetition of gene expression profiling to explore the pathways and mechanisms involved in gonadal develpment and sex determination.